using the LSL-Cas9-Tg model, and that phenotypes observed utilizing these models can be confidently interpreted as being gene specific, and not confounded by the chronic overexpression of Cas9.β-cell microRNA-21 (miR-21) is increased by islet inflammatory stress but it decreases glucose-stimulated insulin secretion (GSIS). Thus, we sought to define the effects of miR-21 on β-cell function using invitro and invivo systems.
We developed a tetracycline-on system of pre-miR-21 induction in clonal β-cells and human islets, along with transgenic zebrafish and mouse models of β-cell-specific pre-miR-21 overexpression.
β-cell miR-21 induction markedly reduced GSIS and led to reductions in transcription factors associated with β-cell identity and increased markers of dedifferentiation, which led us to hypothesize that miR-21 induces β-cell dysfunction by loss of cell identity. In silico analysis identified transforming growth factor-beta 2 (Tgfb2) and Smad family member 2 (Smad2) mRNAs as predicted miR-21 targets associated with the maintenance of β-cell identity. Tgfb2 and Smad2 were confirmed as direct miR-21 targets through RT-PCR, immunoblot, pulldown, and luciferase assays. Invivo zebrafish and mouse models exhibited glucose intolerance, decreased peak GSIS, decreased expression of β-cell identity markers, increased insulin and glucagon co-staining cells, and reduced Tgfb2 and Smad2 expression.
These findings implicate miR-21-mediated reduction of mRNAs specifying β-cell identity as a contributor to β-cell dysfunction by the loss of cellular differentiation.
These findings implicate miR-21-mediated reduction of mRNAs specifying β-cell identity as a contributor to β-cell dysfunction by the loss of cellular differentiation.Injectable hydrogels have been studied as drug delivery systems because of their minimal invasiveness and sustained drug release properties. Pluronic F127, consisting of poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) triblock copolymers, exhibits thermo-responsive properties and hence is injectable due to its rapid sol-gel transition. Unmodified Pluronic F127-based hydrogels, however, have limited long-term stability and controllable release of drugs entrapped within them. In this study, host-guest interactions between adamantane-conjugated Pluronic F127 (F127-Ad) and polymerized β-cyclodextrin (CDP) were employed to develop a hydrogel-based protein delivery system. Single or multiple adamantane units were successfully introduced at the termini of Pluronic F127 with a 100% conversion yield, and the synthesized F127-Ad polymer produced a physically crosslinked micelle-packing structure when mixed with CDP. As the number of adamantanes at the terminal ends of Pluronic F127 increased, the criticthe Pluronic F127-based hydrogel has some limitations in its long-term stability and mechanical property, it is inevitable to modify its structure for the application to drug delivery. In this study, mono- or multi- adamantane-conjugated Pluronic F127s were synthesized and mixed with β-cyclodextrin polymers to form hydrogels with host-guest interaction-mediated micelle-packing structures. The host-guest interaction introduced into the hydrogel system endowed it a sustained protein drug release behavior as well as high durability in vitro and in vivo. By increasing the number of adamantane molecules at the end of the Pluronic F127, both the stability and injectability of the hydrogel could be also modulated.Besides molecular and phenotypic variations observed in cancer cells, intratumoral heterogeneity also occurs in the tumor microenvironment. Correlative stiffness maps of different intratumor locations in breast tumor biopsies show that stiffness increases from core to periphery. However, how different local ECM stiffness regulates key functions of cancer cells in tumor progression remains unclear. Although increased tissue stiffness is an established driver of breast cancer progression, conclusions from 2D cultures do not correspond with newer data from cancer cells in 3D environments. https://www.selleckchem.com/TGF-beta.html Many past studies of breast cancer in 3D culture fail to recapitulate the stiffness of a real breast tumor or the various local stiffnesses present in a tumor microenvironment. In this study, we developed a series of collagen/alginate hybrid hydrogels with adjustable stiffness to match the core, middle, and peripheral zones of a breast tumor. We used this hydrogel system to investigate effects of different local stiffness on moPHOS and fatty acid metabolism responding to stiff matrix microenvironment. The transcriptomic profile of breast cancer cells altered due to microenvironmental stiffness changes.Phloem-feeding insects cause massive losses in agriculture and horticulture. Host plant resistance to phloem-feeding insects is often mediated by changes in phloem composition, which deter insect settling and feeding and decrease viability. Here, we report that rice plant resistance to the phloem-feeding brown planthopper (BPH) is associated with fortification of the sclerenchyma tissue, which is located just beneath the epidermis and a cell layer or two away from the vascular bundle in the rice leaf sheath. We found that BPHs prefer to feed on the smooth and soft region on the surface of rice leaf sheaths called the long-cell block. We identified Bph30 as a rice BPH resistance gene that prevents BPH stylets from reaching the phloem due to the fortified sclerenchyma. Bph30 is strongly expressed in sclerenchyma cells and enhances cellulose and hemicellulose synthesis, making the cell walls stiffer and sclerenchyma thicker. The structurally fortified sclerenchyma is a formidable barrier preventing BPH stylets from penetrating the leaf sheath tissues and arriving at the phloem to feed. Bph30 belongs to a novel gene family, encoding a protein with two leucine-rich domains. Another member of the family, Bph40, also conferred resistance to BPH. Collectively, the fortified sclerenchyma-mediated resistance mechanism revealed in this study expands our understanding of plant-insect interactions and opens a new path for controlling planthoppers in rice.